Potassium (K+) channels are well recognized as a target in treatment of cardiovascular, neurological, renal and metabolic disorders. However, the number of ion channel drugs is strikingly small compared to the number of ion channel genes. Most of the known K+ channel modulators lack potency and specificity. The reason of slow development of K+ channel drugs lies largely in the low capacity and high cost of electrophysiology-based screening techniques. To enable high throughput screening for K+ channel drugs, we developed an assay based on K+ uptake in yeast via the ectopically expressed mammalian inwardly rectifying channel Kir2.1. The assay performed in 96-well plates consists of one "mix and measure" step and is suitable for automation (Z'>0.5). In the screening scheme high throughput yeast-based assay is followed by secondary K+ current measurements in mammalian cells. Thus, in a pilot screen of 10,000 small molecules we identified ~30 compounds that inhibit K+ channels with EC50=0.03-1 [unreadable]M. Even performed on a limited scale, the screening yielded novel modulators that affect both K+ channel activity and cell biology. Based on the validation of the yeast-based assay, we propose to expand the screening using facilities of a Molecular Library Screening Center. Specifically we will: screen >50,000 small molecules for high affinity K+ channel modulators in yeast (Aim 1), and verify primary hits by measurements of K+ current in mammalian cells using 86 Rb+ as a tracer (Aim 2). The goal of the project is to discover both inhibitors and activators of Kir2.1 channel with EC50 < 1 [unreadable]M. Based on our experience, we expect to identify diverse K+ channel modulators with novel mechanisms of action. Identified compounds will serve both for basic study of K+ channel physiology and as a basis for development of clinical K+ channel drugs. [unreadable] [unreadable] [unreadable]